5. Identification of the substrate of MIF and structure of MIF-ligand complexes

6. Study of MIF inhibitors in disease models

7. Co-crystal structures of other protein-inhibitors complexes

Research Summary

We are interested in understanding the mechanism of action of proteins involved in infectious disease, inflammation, and cancer using a variety of techniques including structural biology, yeast genetics, and signal transduction. For example, we determined the three-dimensional structure of two chemokines, human CXCL12 and herpesvirus-8 vMIP-II. We are currently using variety of methods to determine how these proteins interact with theG-protein coupled receptor, CXCR4, a HIV-1 co-receptor and a therapeutic target for cancer metastasis. In collaboration with Demetrios Braddock (Departmentof Pathology), we determined the structure of FIR, a protein that counter-regulates the transcription of the oncogene c-myc. In the area of inflammation we have been studying the signaling mechanism by which macrophage migration inhibitory factor (MIF) exerts its pro-inflammatory effects. We have determined the structure of MIF in the presence of small molecule inhibitors and are studying these inhibitors in mouse models of autoimmunity, inflammation, or infectious diseases.

Extensive Research Description

We are generally interested in understanding the biology, structure, mechanism of action, and pharmacology (inhibition) of proteins that lead to infectious disease, inflammation, and cancer. Our studies are multidisciplinary and include structural biology (X-ray crystallography or NMR), high throughput screening and/or inhibitor design, structure-activity relationships, mutational analysis, the use of a strain of yeast as a model system for chemokines and their receptors, and animal models of disease.

Our current studies include structure-function relationships of chemokines, a family of secreted proteins that recruit immune cells to the site of infection or tissue injury and function by activating specific G-protein coupled receptors. CXCL12 (SDF-1alpha) and vMIP-II/vCCL2 are an agonist and antagonist, respectively, for the GPCR CXCR4, a co-receptor for HIV-1. This receptor also mediates metastasis in a number of cancers, and a C-terminal mutation responsible for an immunosuppressive disease known as WHIM syndrome. Importantly, CXCR4 is the target for Plerixafor, a drug used in peripheral stem cell transplantation. We have determined the structures of CXCL12 and vMIP-II/vCCL2 and are working towards understanding their interactions CXCR4 at high resolution. We are also using random mutagenesis combined with yeast genetics to define smaller peptides with agonist, partial agonist, or antagonist activity.

The other major project is MIF. Inhibition of this "cytokine" MIF by neutralizing antibodies leads to therapeutic efficacy in animal models of sepsis, arthritis, and glomerulonephritis. This protein also inhibits the apoptotic activity of p53. The three-dimensional structure of MIF revealed a striking similarity with two microbial enzymes, including the presence of an enzyme-like crevice with catalytic functional groups. We are now attempting to determine the role of this site in inflammatory diseases and cancer. Interestingly, parasites also express and secrete this protein. In collaborative studies with Drs. Michael Cappello and Richard Bucala, we have determined the structure of MIF expressed from hookworm and Leishmania major, used high throughput screening, and structure-activity relationships to identify compounds with sub-micromolar Ki's.

Other projects include determining the structure of (1) signal transduction proteins involved in inflammation, and (2) enzymes that degrade chemotherapeutic agents.